Genetically defined mouse models have been instrumental in advancing our knowledge of the red blood cell (RBC) and the pathogenesis of inherited anemia. In this proposal, we will continue this powerful phenotype- driven approach to gene discovery and elucidation of mammalian gene function. The specific aims are: Aim 1. Positional cloning of scat and Nan. The recessive mouse mutation, scat (severe combined anemia and thrombocytopenia) and the dominant mutation, Nan (Neonatal anemia), display a severe anemia phenotype resulting from defects intrinsic to hematopoietic stem cells. Genetic mapping localized both scat and Nan to unique positions on chromosome 8 not associated with any previously identified mouse mutation. We hypothesize, therefore, that Nan and scat represent novel genes not previously recognized as significantly impacting blood formation. Here, we will (a) positionally clone the scat and Nan genes, (b) characterize the encoded mRNA and protein products, and (c) further characterize the scat and Nan hematological phenotypes. Aim 2. Characterization of novel mutant sph alleles. Five allelic recessive mouse mutations (spherocytosis, sph) with severe hemolytic anemia due to defects in the 1-spectrin gene (Spna1) have been previously described. We recently identified 2 additional sph alleles, sph3J and sph4J, that display unique phenotypic characteristics compared to the previously described alleles. Hence, we hypothesize that sph3J and sph4J disrupt novel interactions within the RBC membrane skeleton. We will (a) complete the phenotyping of sph3J and sph4J, (b) identify functional deficits incurred by the respective 1-spectrin mutations by characterizing interactions with known membrane skeleton components, and (c) identify novel spectrin interactions within the membrane. Relevance to Public Health: Inherited anemias in mouse and man share common etiologies. Thus, identifying the primary gene defect in mouse models of anemia has direct relevance to human health, will enhance our understanding of blood formation, and provide novel diagnostic and therapeutic targets for hematological pathologies.